Pseudomonas aeruginosa is an opportunistic bacterial pathogen responsible for a significant proportion of serious hospital acquired infections. The organism has intrinsic resistance to most types of antibiotics and can readily acquire resistance to all clinically available antimicrobials, resulting in infections that are impossible to treat. P. aeruginosa colonization occurs in many hospitalized patients, in particular amongst those who have undergone procedures (e.g. intubation or bladder catheterization) that impair local host immune defenses. Some of these patients will develop P. aeruginosa infections, and require aggressive and appropriate antimicrobial therapy to survive; others, however, will not develop invasive disease though P. aeruginosa is repeatedly isolated from respiratory secretions or urine. Although some host factors are correlated with an increased risk of infection, the recent recognition that P. aeruginosa isolates produce distinct complements of virulence factors that are required for virulence in animal models has led several groups to investigate whether such virulence factors also influence clinical P. aeruginosa disease in humans. Such a correlation has indeed been established by two research groups for virulence factors associated with the Type III secretion system of P. aeruginosa; however, no clinically useful test currently exists to measure expression of these virulence factors in patient P. aeruginosa isolates. We propose to develop a rapid diagnostic test to measure virulence factor production by clinical isolates of P. aeruginosa. Phase I specific aims detail the development and optimization of such an assay; Phase II aims are anticipated to include prospective clinical validation of this test. Our hypothesis is that physicians can use information about the virulence factors produced by a given isolate in assessing the likelihood that a patient harboring this isolate will develop serious disease. Such information can improve the care of individual patients, by revealing the presence of highly virulent isolates that warrant aggressive therapy. However, we also believe that such a test may decrease the inappropriate use of broad-spectrum antibiotics in patients carrying avirulent organisms and showing no overt signs of infection. This can have significant consequences both for public health, by limiting the emergence of multi-drug resistant pathogens in our hospitals, and for containing the cost of medical care.